Method and apparatus for sensing ion concentrations in a fluid sample

ABSTRACT

The invention provides a method for the measurement of a concentration of a charged species in a sample, the sample having a plurality of types of charged species and at least one insoluble component. The method comprises: providing the sample on a surface of a partly permeable layer; allowing components of the sample to pass through the partly permeable layer into a channel; and separating the components into sections, such that each at least one of the sections substantially comprises a single type of the plurality of the types of charged species, and determining the charge concentration in the at least one of the sections.

PRIORITY APPLICATIONS

This application is continuation of U.S. patent application Ser. No.12/515,635 filed May 20, 2009 which is a 371 application ofInternational Application No. PCT/EP2006/011148 filed Nov. 21, 2006. Theentire disclosure of each of the foregoing applications is herebyincorporated herein by reference.

FIELD OF THE INVENTION

The invention relates to sensors of charged species in biological,chemical, industrial or environmental samples. In particular, theinvention relates to a method and a sensor for measuring charged speciesconcentrations, in particular ion concentrations, for example lithiumion concentrations, in samples, such as blood. The invention alsorelates to a method for the production of such a sensor.

BACKGROUND AND RELATED ART

Inorganic ions are an essential requirement for life and are found inlarge amounts in drinking water, blood and every cell of an organism aswell as in the environment. For example, the concentration of many ionsi.e. sodium, potassium, magnesium, and calcium inside and outside ofcells is essential for any living organism. Consequently, the ionconcentration in the blood and blood cells of animals and human beingsalso is of high importance for a large variety of body functions.

Normally lithium is a trace element present in the blood plasma, but itis used as a drug to treat bipolar mood disorder. It is estimated thatworldwide over one million people take lithium on a daily basis. Adisadvantage in the use of lithium is the very low therapeutic index,i.e., the ratio between the toxic concentration and the therapeuticconcentration. Most patients respond well to a blood plasmaconcentration of 0.4-1.2 mmol/L lithium while toxic effects can occur ata lithium concentration of above 1.6 mmol/L. A prolonged high bloodlithium level can even result in permanent damage to the nervous systemand even death. Monitoring of the lithium concentration during treatmentis therefore essential, with regular checks every couple of months tokeep the lithium level at desired level.

To avoid extensive operator handling, ion-selective electrodes (ISEs)are routinely used for measurements of blood parameters in an automatedfashion. These ISEs are fast and offer a large dynamic range; however,their response is logarithmic and the required high selectivity forlithium can be a problem. Additionally, in case of lithium intoxicationa fast procedure for blood analysis is required. Currently, a venousblood sample must be withdrawn from the patient by specially trainedpersonnel and transported to the central laboratory and the blood cellsneed to be removed before the measurement is made. This procedure cantake up to 45 minutes. To minimize sample throughput time and enablemeasurements on location, miniaturized devices employing ion-sensitivefield-effect transistors are available to determine the concentration ofpotassium and sodium in whole blood even as a hand-held analyzer.However, such analyzers are not used for lithium determination, becauseof the high background concentration of other charged species, inparticular sodium ions, compared to the much smaller concentration oflithium ions.

The direct measurement of lithium in whole blood and the determinationof inorganic cations in blood plasma have been described anddemonstrated by E. Vrouwe et al. in Electrophoresis 2004, 25, 1660-1667and in Electrophoresis 2005, 26, 3032-3042. Using microchip capillaryelectrophoresis (CE) with defined sample loading and applying theprinciples of column coupling, alkali metals were determined in a dropof whole blood. Blood collected from a finger stick was transferred ontothe chip without extraction or removal of components. The lithiumconcentration can be determined in the blood plasma from a patient onlithium therapy without sample pre-treatment. Using a microchip withconductivity detection, a detection limit of 0.1 mmol/L has beenobtained for lithium in a 140 mmol/L sodium matrix.

In these disclosures, the components of the blood sample are separatedelectrophoretically inside a micro-channel. A double T injectiongeometry is used to select the ion components of interest and to guidethem to detection electrodes.

In these systems, the sampling loading has to be well defined in orderto ensure the correct separation of blood plasma components in thedouble T geometry. In addition, the double T geometry is complicated toapply and not well suited for easy to use applications.

SUMMARY OF THE INVENTION

The invention provides a method for the measurement of a concentrationof a charged species in a sample, the sample having a plurality of typesof charged species and at least one insoluble component, the methodcomprising: providing the sample on a surface of a partly permeablelayer; allowing components of the sample to pass through the partlypermeable layer into a channel; and separating the components intosections, such that each at least one of the sections substantiallycomprises a single type of the plurality of the types of chargedspecies, and determining the charge concentration in the at least one ofthe sections.

Thus, the invention provides a method for dividing a sample, inparticular a biological sample such as blood plasma into sections, eachsection comprising substantially one or a one group of charged speciesand subsequently determining the concentration of charged species inthis section.

The invention also provides an apparatus for the measurement of aconcentration of a charged species in a sample, the sample comprising aplurality of types of charged species and at least one insolublecomponent, the apparatus comprising at least one channel with at leastone opening, a partly permeable layer covering the at least one opening,at least two electrophoresis electrodes arranged along the at least onechannel on each side of the opening, and at least one sensor formeasuring at least one type of charged species in the at least onechannel.

The method and the apparatus are particularly useful for the measurementof ion concentrations of biological samples, for example blood plasma.The ions measured include but are not limited to sodium, potassiummagnesium, calcium and the like. In one application of the invention,the sample may also contain lithium. In this case, the preferred ion tobe measured is lithium but may be any other ion present in the sample.The invention is equally applicable to other charged species such aslipids, DNA or other polyelectrolytes or electric charge carryingpolymers.

The concentration of a first one of the plurality of type of chargedspecies may be determined relative to a second one of the plurality ofthe types of charged species. The first type of charged species may belithium ions and the second type of charged species may be sodium ions;thus the ratio between lithium and sodium ions in the sample can bedetermined.

The at least one channel may have a single opening covered by apartially permeable layer. Using the single opening for sampleapplication, electro-osmotic pressure or hydrodynamic pressure and anyhydrodynamic flow inside the channel can be advantageously avoided. Inthat way, diffusion is the main or only transport mechanism.

In one embodiment, the at least one channel may have two openings in theotherwise sealed channel system. Using hydrodynamic pressure sampleinjection is realized by convective flow form one opening towards theother. In this specific case one opening is covered with the samplewhile the other opening is not.

The partially permeable layer may be a membrane separating the samplefrom the at least one channel. The membrane may be permeable to ions orother charged species while the membrane may be impermeable to largercomponents. In particular, the membrane may be impermeable to theinsoluble component. The membrane may also be a gas-permeable membranethat is impermeable to liquids. The partially permeable layer may be aseparate layer placed on top or below the at least one opening of thefirst cover layer.

A membrane holder may be used on the first cover layer for placing themembrane on the first cover layer. The membrane holder may be attached,i.e. glued the first cover layer or formed directly in the first coverlayer.

The permeable layer may also be a region of the first cover layer thatis made partially permeable. The permeable layer may comprise at leastone region with a hydrophilic surface. Additionally, the permeable layeror the first cover layer may comprise at least one region with ahydrophobic surface.

The permeable layer may also consist of one or more holes in thechannel. The sample may thus come into direct contact with a solutioninside the channel.

The sample also comprises at least one insoluble component, i.e. in thecase of a biological sample such as blood, red blood cells, white bloodcells, platelets and the like that are usually present in the blood.Thus the present invention advantageously allows for the determinationof an ion concentration in whole blood without prior purification ortreatment thus avoiding any laboratory pre-treatment of the sample. Theinvention is therefore particularly useful for the application inpatient operated system that do not require a specially trainedphysician or medical care taker.

The at least one sensor comprises one or more pairs of conductivityelectrodes for determining the charge concentration in the at least oneof the sections substantially comprising the single type of theplurality of the types of charged species. For example, a first pair ofconductivity electrodes may be arranged in or nearby the channel at somedistance from the at least one opening for measuring the concentrationof charged species of a first polarity. A second pair of conductivityelectrodes may be arranged at the opposite end of the channel fordetermining the concentration of a second charged species of oppositepolarity to the first polarity.

The invention also provides a method for the manufacture of an apparatusfor measuring the concentration of charged species in a sample, themethod comprising providing a substrate, forming a channel into thesubstrate, placing a first cover layer on the substrate, such that thefirst cover layer covers the channel, whereby the first cover layercomprises at least one opening providing access to the channel, andplacing a partly permeable layer on the at least one opening.

Using this method for the production of the apparatus, the partlypermeable layer may be placed on the at least one opening prior to,after or simultaneously with placing the first cover layer on thesubstrate.

Prior to use of the apparatus, the at least one channel may be filledwith an electrolyte. In one embodiment the filling of the channelcomprises evacuating air and sucking electrolyte into the channel. Theelectrolyte may be filled into the at least one channel prior tocovering the channel with a second cover layer.

DESCRIPTION OF THE DRAWINGS

The invention may be better understood with respect to the figures andthe detailed description of preferred embodiments, which is illustrativeonly and not limiting to the invention and wherein:

FIGS. 1 a to 1 d show main components of an apparatus according to theinvention in a top view and FIG. 1 e shows a side view of the componentsof FIGS. 1 a to 1 d assembled to an apparatus according to theinvention.

FIG. 2 shows a section of FIG. 1 e in greater detail

FIGS. 3 a to 3 f show main steps for providing a sample to be measuredto the micro channel in the enlarged and detailed view of FIG. 2.

FIGS. 4 a and 4 b show an example of an apparatus according to theinvention in top view and in side view, respectively, FIG. 4 c and FIG.4 d show electrode configurations for conductivity detection, both,contactless (FIG. 4 c) and in contact conductivity detection (FIG. 4 cand FIG. 4 d) are possible realizations. FIG. 4 e shows two possiblebackground measurement signals at for example two different measurementtemperatures.

FIGS. 5 a and 5 b show alternative embodiments of the present inventionand FIG. 5 c shows examples of corresponding measurement signals.

FIG. 6 a shows another embodiment of the apparatus with a substantiallyU-shaped channel.

FIG. 6 b shows a further embodiment with two opening in a singlechannel.

FIG. 7 shows a further embodiment of the invention with a membraneholder.

FIGS. 8 a and 8 b show an embodiment of the invention with an extraelectrode.

FIGS. 9 a to 9 d illustrate a method of the invention in which the fluidis inserted into the channel by vacuum.

FIG. 10 shows a further embodiment of the invention in which the fluidis inserted into the channel by use of second opening in the channel.

In the figures same reference numerals describe the same or similarobjects.

DETAILED DESCRIPTION OF THE INVENTION

FIGS. 1 a to 1 d show the components of an apparatus according to theinvention in a top view.

The apparatus comprises a substrate 10 into which a channel 12 isformed, as shown in FIG. 1 a. The substrate 10 may be made from glass orplastics material. Any other material allowing for the fabrication ofchannels 12 may be used. In case of glass as the substrate material, thechannel 12 is etched into the substrate 10 between a first reservoir 14and a second reservoir 16 and the side walls of the channel 12 arecoated with a polymer. The channel 12 may be of sub-centimetredimensions, in particular the channel 12 may be less than 1 cm in widthand less than 100 μm in depth. The first reservoir 14 and the secondreservoir 16 may be considerably larger in size than the width of thechannel 12 (e.g. 100 μm to 1 cm), but may have substantially the samedepth. The channel 12 and the first reservoir 14 and the secondreservoir 16 may be filled with an electrolyte prior to use. This can bedone, for example, by evacuating the channel 12, the first reservoir 14and the second reservoir 16 and then allowing the electrolyte to besucked into the channel 12 and the first reservoir 14 and the secondreservoir 16. The first reservoir 14 and the second reservoir 16 can forexample serve for equilibrating pressure differences to ensure that thechannel 12 is always filled with the electrolyte.

The channel 12 may also be made of a plurality of nanochannels having awidth of between 1 and 500 nm. The small size of the nanochannelssuppresses hydrodynamic and electro-osmotic pressure within the channel12.

The apparatus further comprises a first layer 20 shown in FIG. 1 b as acover layer for covering in use the substrate 10 and for closing thechannel 12 to prevent in use any fluid like the electrolyte and thesample inside the channel 12 from evaporation or leaking out of thechannel 12. The first layer 20 may be made for example, from glass, apolypropylene film or hydrophobic membrane, such as those supplied bythe Pall Corporation under the designation Supor Membrane Disk Fillers(hydrophilic polyether sulfone) or Millipore Durapor(polyvinylidene—PVDE) and may have a thickness of less than 1 mm, inparticular less than 1 μm. The first layer 20 is non permeable. Thefirst layer 20 provides a first opening 22 to be arranged on top of thechannel 12 in order to provide access for the sample to the channel 12.The access opening 22 may have the form of a circle but any formsuitable for inserting liquid into the channel may be used.

In addition, according to the invention a membrane 30 is provided, shownin FIG. 3 c. In the example shown, the membrane 30 is in use arranged ontop or below of the opening 22 of the first layer 20. The membrane 30may be made of a permeable hydrophilic and/or biocompatible polymer of 1to 100 μm thickness that is semipermeable, for example, nitrocellulose.It is possible that the membrane 30 be placed on the channel 12 prior tothe first layer 20. Thus the membrane 30 may also be arranged betweenthe first layer 20 and the substrate 10. The membrane 30 may also beintegrated into the first layer 20. In any case, the membrane 30 ishydrophilic and can be made, for example, from nitrocellulose.

The size and the properties of the membrane 30 may be adapted to allowfor diffusion of species or transfer of a specified volume of a samplefrom the sample side to the inside of the channel 12 in order to enablecomparable measurements.

According to one aspect of the invention, the membrane 30 is permeableto blood plasma and its components in the sample but filters out largerinsoluble components such as cell material in the sample or the like. Inthis way, cell material like red blood cells, white blood cells,platelets or the like are filtered out and only blood plasma enters thechannel 12 for further examination. Other components may also befiltered out.

According to another aspect of the invention, the membrane 30 ispermeable to charged species inside the blood plasma and the membrane 30covered first opening 22 is the only opening to the channel. It may alsobe the only opening enabling convective flow into the channel 12. Inthat way convective flow is suppressed and at least the blood plasma andall kinds of cell material are prevented from entering the channel whileonly the charged species, in particular the ions diffuse into thechannel 12 for further examination.

In a further embodiment of the invention, the membrane 30 and the firstlayer 20 might be made in a single step in which the first layer 20 is apolymer film which is made to act locally as a membrane or the firstlayer 20 is a polymer film in which the full polymer film is a membranein which the hydrophobicity is altered. In the latter case, thehydrophobicity of the film is changed such that the film is hydrophilicat the position at which the sample is to be injected.

More than one access opening 22 may be made in the first layer 20. Thisis useful, for example, for allowing the sample to enter into thechannel 12 at multiple entry points. This allows for multiplemeasurements to be made and averages to be taken. One further advantageof more than one access opening 22 is to allow convective flow from oneopening towards another opening and thus providing an alternativetransport mechanism through the opening 22 into the channel 12.

The membrane 30 can also be provided with microneedles on its surface topuncture the skin to obtain the sample more easily. Furthermore themembrane 30 could itself be punctured to realize, alter or improve itsporosity.

A second polymer film 40 shown in FIG. 1 d is provided for covering thefirst layer 20 and the semipermeable membrane 30 in order to protect thefirst layer 20 and the semipermeable membrane 30 from contamination, tokeep them sterile and/or clean prior to use and to prevent leakage offluid from the channel. Should the semipermeable membrane 30 havemicroneedles, these microneedles are also protected by the secondpolymer film 40. The second polymer film 40 is made of, for example,polypropylene. The second polymer film 40 may be removed immediatelyprior to use and a blood sample, i.e. a droplet of whole blood may inuse be placed on top of the semipermeable membrane 30. The secondpolymer film 40 may have a loose end so that it can be easily gripped tobe removed prior to use of the apparatus 2.

FIG. 1 e shows a side view of the components of FIGS. 1 a to 1 dassembled as an apparatus 2 according to the invention. The first layer20 is placed in top of the substrate 10 thus covering the top side ofthe channel 12. The first layer 20 has an opening 22 arranged on top ofthe channel 12. The opening 22 is covered by the membrane 30. In thecase shown in FIG. 1 d the apparatus 2 is covered by the second polymerlayer 40 covering the whole or part of surface of the apparatus 2 andthus protecting the apparatus 2 from damage, dust, evaporation, etc.

The first layer 20 may also include hydrophobic membranes permeable togas. The function of the gas permeable hydrophobic membrane is toprevent over pressure which might build up in the channel 12 as will beexplained later. The gas permeable hydrophobic membrane might be appliedseparately but also embedded in the first layer 20.

FIG. 2 shows an exploded view of the area marked by a circle in FIG. 1 ein greater detail. The membrane 30 is placed on top of the opening 22 inthe first layer 20. The first layer 20 covers the channel 12 in thesubstrate 10 leaving an access to the channel 12 via opening 22. Theopening 22 is covered by the membrane 30, thus, in use, only componentsthat can diffuse or pass otherwise through the membrane 30 can accessthe channel 12. For protection and for preventing unwanted access to orcontamination of the membrane 30, the membrane 30 is covered by a secondpolymer film 40. The membrane may be glued or otherwise fixed on, underor in the first layer 20. It would be possible to mount the membrane 30in a holder and insert this holder in the opening 22 of the first layer20. An example of a holder is described below with respect to FIG. 7.

The channel 12 may be coated with polymers in order to suppress electroosmosis flow as is known in the art.

FIGS. 3 a to 3 f show the main steps for providing the sample to bemeasured to the channel 12 in the enlarged and detailed view of FIG. 2.

FIG. 3 a illustrates a detailed view of the apparatus of FIG. 2, wherebythe channel 12, the opening 22 and the membrane 30 are filled with abackground solution (shown as grey areas in the Fig.). For the detectionof lithium, the background solution can be a background electrolyte(BGE) solution containing for example 50 mmol/L2-(N-morpholino)ethanesulfonic acid and 50 mmol/L histidine at pH 6.1.Glucose may be added, for example about 200 mmol/L for adjusting theosmotic strength of the background solution. Other background solutionsmay be used depending on the charged spieces, i.e. the ion to bemeasured. The second polymer film 40 protects the apparatus 2 and thesolution and prevents the solution from being contaminated prior to use.FIG. 3 a illustrates the form in which the apparatus 2 may be shipped toa user.

FIG. 3 b shows the removal of the second polymer film 40 prior to use ofthe apparatus 2. The second polymer film 40 serves as a protecting layerfor protecting the membrane 30 and the first polymer layer 20 duringshipping and storage of the apparatus 2. As shown in FIG. 3 b, thesecond polymer film 40 is removed from the apparatus 2 in order toprovide access for the sample to the membrane 30. The second polymerfilm 40 is provided with a quick release mechanism, such as a pull-tab,to allow easy removal of at least part of the second polymer film 40.

Prior to placing the sample on the membrane illustrated in FIG. 3 c, oneor more apparatus parameters, such as the conductivity of theelectrolyte or temperature may be measured, for calibration or as asystem check. A conductivity measurement of the pure electrolyte mayalso be performed as a system check, i.e. to check that electrolyte ispresent in the channels and that the measurement system is workingcorrectly. It is advisable to flush the channel 12 electrokineticallyprior to carrying out the measurement. This is to get rid of the firstdiffused parts of the sample in the channel 12. The conductivitymeasurement might be used for temperature measurements. The conductivitymeasurement might also be used as an internal check of the condition ofthe apparatus 2. The later might be realized with another temperaturemeasurement method implemented somewhere in or around apparatus 2.

Heating elements may be placed inside or around the channel 12 or aroundthe apparatus to alter the temperature of the liquid in the channel 12.The change of conductivity as a function of temperature may be used forcontrol or calibration.

In FIG. 3 c a sample 50, i.e. an untreated whole blood sample, is placedon the upper surface of the membrane 30. The membrane 30 is hydrophilicand permeable. Thus the sample 50 will be absorbed and pass through themembrane 30 as shown in FIG. 3 d, whereby cell material such as redblood cells, white blood cells, etc are filtered out. This is done asthe cell material might break down inside the channel 12 and alter theconcentration inside the channel 12. The size of the pores of themembrane 30 might also be adjusted to filter out, for example, lipids orother larger components so that only electrolytes pass into the channel12. Diffusing through the membrane 30, the filtered sample 50 will comein contact with the first layer 20 and enter into the opening 22.

As illustrated in FIGS. 3 d and 3 e, the filtered sample 50 diffusesthrough the opening 22 into channel 12 of the substrate 10. The amountof the filtered sample 50 reaching the channel 12 is determined by thesize of the opening 22, the properties of the membrane 30, theproperties of the sample 50 as well as the electrolyte present inchannel 12.

FIG. 3 f illustrates how a portion of the filtered sample 50 thatdiffused into the channel is electrophoretically separated in thechannel 12 when an electrical field is applied along the channel 12. Theelectrical field will separate all of the charged species in thefiltered sample and move the charged species towards the reservoirs 14and 16 at the end of the channel 12.

Electrodes for providing an electrical field along the channel 12 may beimbedded or inserted in the first reservoir 14 and the second reservoir16. It is also possible that a plurality of electrodes are placed alongthe channel 12 to create extra strong fields at those locations wherethe separation of the ions is necessary, by switching the electric fieldfrom one area to another. It was explained above that gas permeablehydrophobic membranes are used in the apparatus to prevent overpressureoff gas. This overpressure may occur at the electrodes because ofelectrolysis.

The measurement may be performed repeatedly.

FIGS. 4 a and 4 b show an example of an apparatus 2 according to theinvention in top view and in side view, respectively, wherein the firstreservoir 14 comprises a first electrophoresis electrode 64 and thesecond reservoir 16 comprises a second electrophoresis electrode 66. Byapplying an electrical voltage to the electrophoresis electrodes 64, 66,charged particles inside the channel 12 may be separated or moved alongthe channel 12. The electrophoresis electrodes 64, 66 may be made of anyconducting material. Examples of electrodes used include, but are notlimited to, titanium electrodes with a chrome layer or silver/silverchloride electrodes The electrophoresis electrodes 64, 66 can beintegrated in the substrate 10 or may be otherwise mounted into thereservoirs 14 and 16 or any other place in channel 12.

In an alternative embodiment, the electrophoresis electrodes 64, 66and/or the conductivity electrodes 72, 74 may be mounted to ameasurement device on which the apparatus 2 can be mounted formeasurement.

The electrodes 72, 74 are not limited to a solely two-way electrodearrangement but can exist of multiple electrode arrangement.

A voltage may be applied to the electrophoresis electrodes 64, 66 by apower supply or any means known in the art.

FIG. 4 c shows an exploded top view of the area marked by a circle inFIG. 4 a and FIG. 4 d shows an exploded side view of the same area in aside view, as also marked by a circle in FIG. 4 b. In this area twoconductivity electrodes 72 and 74 are provided in close proximity to orinside the channel 12 for measuring the conductivity of the fluid acrossthe channel 12 at the position of the conductivity electrodes 72, 74.The conductivity electrodes 72 and 74 may be integrated in the substrate10 and at least partially extend into the channel 12. As shown in FIG. 4d, the conductivity electrodes 72, 74 may be arranged on the bottom ofthe channel 12 but any other position at the channel 12 is possible. Theconductivity electrodes 72, 74 may be connected to conductivitymeasurement known in the art.

In one embodiment of the invention, two pairs of conductivity electrodes72 and 74 are used. One of the pairs of conductivity electrodes measurespositive ions and the other one of the pairs of conductivity electrodesmeasures negative ions in the channel 12. The two pairs of conductivityelectrodes 72 and 74 are placed on either side of the opening 22 throughwhich the sample enters into the channel 12.

Placement of the conductivity electrodes 72 and 74 as well as theelectrophoresis electrodes 64, 66 may be carried out during or after themanufacture of the apparatus 2. For example, the conductivity electrodes72 and 74 and the electrophoresis electrodes 64, 66 may be pushedthrough the surface of the polymer cover 20 or the substrate 10 into thechannel 12; thus costly implementation of the conductivity electrodes 72and 74 and the electrophoresis electrodes 64, 66 in the chip can beavoided.

The conductivity in the channel 12 between conductivity electrodes 72and 74 can be monitored over time. In case no charged component or anequal distribution of charged particles is present inside the channel,for example the BGE solution, a constant or relatively slowly varyingconductivity will be measured and monitored as illustrated in FIG. 4 e.

In case of the insertion of charged species, such as ions or the like,into the channel 12 using the method described with respect to FIG. 3,the charged species are moved along the channel 12 by an electric fieldapplied between the electrophoresis electrodes 64 and 66. The chargedspecies will be separated electrophoretically while travelling along thechannel 12. For example, Na-ions of a blood sample 50 will move fasterthan Li-ion that may also be present in the blood sample 50. Thus, twopeaks will be measured consecutively by the conductivity electrodes 72and 74. A first peak represents the faster moving Na-ions passing theconductivity electrodes 72 and 74 and a second peak represents theslower moving Li-ions passing the conductivity electrodes 72 and 74. Itis obvious to the person skilled in the art that more than two types ofions can be measured and that any charged component that may beseparated by electrophoresis means can be monitored in that way.

The invention may be applied to measure absolute ion concentrations orfor the measurement of relative ion concentrations, i.e. for themeasurement of Na/Li-concentration ratios.

Further measurement electrodes or other types of sensors, i.e. opticalsensors such as fluorescence sensors as known in the art may be added tomeasure the concentration or presence of further species in the samplewithin the same measurement. Capacitative sensors can also be used.

Prior to the measurement of the concentration of the charged species,such as ions, it is useful to measure the conductivity of theelectrolyte in combination with the temperature of the apparatus toensure that apparatus is performing correctly.

FIGS. 5 a and 5 b show alternative embodiments of the present invention.These embodiments may for example be used for calibration purposes.

FIG. 5 a shows an apparatus 102 according to the invention and based onthe apparatus 2 described above. In this embodiment of the invention achannel 112 between a first reservoir 114 and a second reservoir 116branches into a first channel branch 111 and a second channel branch113. Both the first channel branch 111 and the second channel branch 113of the channel 112 are reunited before the second reservoir 116. Thefirst channel branch 111 is considerably longer than the second channelbranch 113. Both the first channel branch 111 and the second channelbranch 113 have an opening 122 and 123, respectively. The openings 122and 123 are each covered with a membrane 130 and 131, respectively.

If two different samples 150 and 151 are each placed on separate ones ofthe membranes 130 and 131 and an electric field is applied along thechannel 112, the ions of each of the samples will be separated and movedalong the channel 112. As the first channel branch 111 is longer thanthe second channel branch 113, the charged species, i.e. ions, of thesecond sample 151 will arrive first at channel 112 while the chargedspecies of the first sample 150 take somewhat longer. Thus both of thecharged species can be measured independently one after the other withthe same pair of conductivity electrodes (not shown) resulting in asignal as illustrated in the top line of FIG. 5 c.

This embodiment may also be used for calibration by providing a knownsample 150 to membrane 130 resulting in a corresponding first signalthat can be used for calibration. The second signal from an unknownsample 151 provided to membrane 131 will arrive later in time due to thelonger channel branch 111. The strength of the second signal can than becompared to the first calibration signal and the concentration of thecharged components in the unknown sample can be determined as known inthe art.

This embodiment might also be used with same sample provided to membrane130 and to membrane 131 to realize higher accuracy by for instanceaveraging.

FIG. 5 b shows an alternative embodiment of the invention where twochannels 212 and 213 are arranged in parallel. Each of the channels 212and 213 are basically identical to the embodiment of FIGS. 1-4 with theadvantage that two samples 250 and 251 are placed in parallel onmembranes 230 and 231, respectively, so that both samples are measuredin parallel. As both channels 212 and 213 are identical, themeasurements can be compared. Examples are shown in the lower lines ofFIG. 5 c

For calibration purposes, one sample, for example a first sample 250 canbe a known sample with known ion concentrations. Thus the signal offirst sample 250 can be used for calibration and compared to a signalfrom a second sample 251 and second channel 213 and the concentration ofcharged particles can be determined in a way known in the art.

It is obvious, that a plurality of channels can be arranged in parallel,for example to perform multiple measurements to accelerate throughput orto increase measurement statistics.

FIG. 6 a shows yet another embodiment of an apparatus for themeasurement of a concentration of an ion in a sample wherein a channel312 is substantially curved and a first reservoir 314 comprising a firstelectrophoresis electrode 364 is place at the same side of a substrateas a second reservoir 316 comprising a second electrophoresis electrode366. Contacts for both of the electrophoresis electrodes 364 and 366 maybe guided to the side of the apparatus for easy contact to the side ofthe apparatus. In addition the conductivity electrodes 372 and 374 areprovided in proximity to the second reservoir 366 for measuring theconductivity of charged component in the channel 312 at this position.The conductivity electrodes 372 and 374 may connected via contacts thatare arranged at the same side of the apparatus or substrate as thecontacts for the conductivity electrodes. In that way, only the part ofthe apparatus with the contacts needs to be placed into contact with ameasurement device and free access to the membrane 330 placed in opening322 can be ensured. With such an apparatus it is possible to have easyaccess, for example with a finger tip to the membrane 330, while theapparatus is inserted or contacted to a measurement and/or controldevice. The channel 312 is further straight between the opening 322 andthe conductivity electrodes 372, 374 so that no bending of the channel312 containing the sample is necessary which might influence measurementaccuracy or make measurement otherwise difficult.

FIG. 6 b shows a modification of the embodiment shown in FIG. 6 a,further providing a second opening 423 in channel 412 that is covered bythe same membrane 430 as a first opening 422. Thus a sample on membrane430 will diffuse substantially in the same time through both of theopenings 422 and 423 in the channel 412. Applying an electrical field toelectrophoresis electrodes 464 and 466, will, depending on the sign ofthe voltage, cause for example the positively charged species or ions tomove into a first channel section 411 towards a second electrophoresiselectrode 466. Similarly negatively charged species are moved into achannel section 412 towards a first electrophoresis electrode 464. Theconductivity electrodes 472, 474 and 471, 473 allow for measurement ofboth of the positively charged species and the negatively chargedspecies. Thus the charged species of both electrical charges can bemeasured in parallel.

FIG. 7 shows a modification of the apparatus according to the inventionshown in FIG. 2. A membrane holder 32 is mounted on top of the secondlayer 20. The membrane 30 is mounted, for example glued, onto or in themembrane holder 32. Thus the membrane can be assembled on the membraneholder before the membrane holder is mounted on the apparatus.

The membrane holder 32 may be made from plastics material.

In the embodiment shown the membrane holder 32 forms a “cup”-like or aring like structure providing a receiving section for the membrane 30.The upper surface of the membrane is substantially planar with the upperrim of the “cup”-like structure of the membrane holder. The membraneholder provides thus a frame for the membrane 30 with a well definedsurface area of the membrane being left for contact with the sample. Inthat way, the amount of sample coming in contact with the membrane canbe controlled in a simple and efficient way, even when the sample ismuch bigger, than the membrane.

The walls of the membrane holder may also be higher than the thicknessof the membrane, thereby providing a “cup”-like or ring-like structurefor the sample (not shown) with the membrane at the bottom of the “cup”.The cup may be used to collect the sample on the membrane.

The membrane holder 32 may enable a fast and easy exchange orreplacement of the membrane 30. By exchanging the membrane 30, theapparatus can be easily adapted to different measurements, e.g. by usingmembranes with different pore sizes, the size of components that arefiltered out or let into the channel can be adjusted to the needs of theparticular measurement.

The membrane holder 32 can furthermore enable easy fixation of themembrane 30 on the first cover layer by for instance a click-and-fixmethod.

The membrane holder 32 can have the second cover layer 40 on top toprevent leakage, evaporation, etcetera.

FIGS. 8 a and 8 b show an embodiment of the present invention with anadditional anti-tailing electrode 65 for preventing tailing of thesample or components inside the channel 12. The anti-tailing electrode65 is shown in between first cover layer 20 and membrane 30. Theanti-tailing electrode 65 may, however, also be arranged differently onthe top side of or at the opening 22 of first cover layer 20. FIG. 8 ashows the apparatus with the anti tailing electrode 65 in the same stateas the apparatus shown and described with respect to FIG. 3 e. Theapparatus and the method described with respect to FIGS. 1 to 3 applyaccordingly and the filtered sample may thus diffuse through membrane 30and first opening 22 into channel 12 as described above.

Prior or simultaneously to applying the electrical field along thechannel 12 for electrophoretically separating the portion of thefiltered sample illustrated and described with respect to FIG. 3 f,voltage is applied additionally to anti-tailing electrode 65. Thereby aportion of the sample component is also driven backwards through thefirst opening 22 towards the membrane 30 as indicated by arrow 800 inFIG. 8 b. The electrical field separates the charged species in thefiltered sample and move the charged species towards the reservoirs 14and 16 at the end of the channel 12 and towards the membrane 30.Therefore, no sample component enters the channel after startingseperation. This effect increases measurement accuracy.

The extra electrode 65 may also consist of a plurality of electrodes andmight also be used for parameter detection prior or during measurement.

FIGS. 9 a to 9 d show how a fluid such as the background electrolytesolution (BGE) or any other solution may be inserted into the channel 12of the apparatus described with respect to FIGS. 1, to 3 using only oneopening 22 in the channel 12. FIG. 9 a illustrates the apparatus of FIG.2 before any fluid is inserted. A droplet of fluid 14 is put on themembrane 30 as shown in FIG. 9 b. The fluid 14 will then flow into themembrane 30 until it covers opening 22 of channel 12. At this point,illustrated in FIG. 9 c, the fluid does not enter by itself further intothe channel 12 because of the air or gas being inside the channel 12.The air of gas inside the channel 12 can only exit the channel 12through the single opening 22, which is covered by fluid 14. FIG. 9 dshows that by the application of a vacuum (indicated by arrow 900) theair or gas inside the channel 12 can be sucked out of the so that thefluid 14 enters into the channel 12.

FIG. 10 shows a further method for sampling a fluid such as blood or anyother sample into the micro-channel 12. A second opening 23 may beprovided at some distance of first opening 22. Both openings 22 and 23are connected by the channel 12. Preferably, the channel 12 has nofurther openings that said openings 22 and 23 and is otherwise sealed.The second opening 23, however, is not covered by a membrane. Fluidinside the channel 12 may exit through the second opening 23, when thesample is applied on the first opening 22.

The second opening 23 may be covered by a polymer layer or otherwiseclosed, after the fluid has been filled into the channel 12, to preventevaporation of the fluid. During sampling the second opening 23 has tobe connected in any way to air and might not be covered by the sampledirectly.

Connections to the electrodes can be also arranged on one side of theapparatus allowing for easy attachment and connection to a measurementdevice. Easy access is especially important when the apparatus is inform of a disposable chip that can be inserted for one measurement intoa measurement device that may be operated by the patient.

The apparatus 2 can be packaged inside a packaging with suitableinterfaces to allow connection to electronics for measurement andcontrols, communications interfaces and display interfaces as well asfor power electronics.

The openings 22 have been described as being made in the upper surfaceof the substrate 10. However, the openings 22 can also be realized atany other location of the apparatus 2 for instance in the side.

The apparatus 2 can be easily used by a patient to measure theconcentration of ions in blood. For example, for those patientssuffering from bipolar mood disorder, the patient can measure theconcentration of lithium ions in the blood on a regular basis. Shouldthe concentration go below a critical level (e.g. 0.4 mmol/L) then thepatient can take extra lithium. Should the concentration go above acritical level (1.0 mmol/L), then the patient can stop or lowermedication and if necessary be hospitalised.

The use of the apparatus 2 has been described with respect to themeasurement of lithium ions. The apparatus 2 could also be used for themeasurement of potassium and/or phosphate ions to observe thefunctioning of a kidney or sodium and/or potassium ions to determinedehydration.

The apparatus of the invention has applications outside of the medicalfield. For example, it would be desirable when using the apparatus inthe environmental and other fields to be able to use the same apparatusover the course of a period of time. In this case, the apparatus mightbe provided with a plurality of openings 22, each of which had its owncover. The own cover would be periodically removed from different onesof the plurality of openings 22 to allow repeated measurements.

The invention has been described with respect to several embodiments. Itwill, however, be clear to those skilled in the art that the inventionis not limited thereto. Rather the cope of the invention is to beinterpreted in conjunction with the following claims.

1. A method for conducting a measurement on a sample using a disposableapparatus, the method comprising the steps of: removing a cover layerfrom a partly permeable layer comprising one or more holes providingfluidic access to at least one channel, the at least one channel beingfilled with a background solution and being otherwise sealed, providinga sample onto a surface of the partly permeable layer, allowingcomponents of the sample to pass through the partly permeable layer intothe at least one channel, determining at least one property of thecomponents of the sample.
 2. The method according to claim 1, whereinthe providing of the sample comprises obtaining the sample withoutpre-treatment from the human body.
 3. The method according to claim 1,wherein the components of the sample comprise a plurality of types ofcharged species.
 4. The method according to claim 3, further comprisingchecking the presence of the components of the sample in the at leastone channel after the allowing of the components of the sample to passthrough the partly permeable layer into the at least one channel.
 5. Themethod according to claim 1, wherein the determining of the at least oneproperty of the components comprises a conductivity measurement.
 6. Themethod according to claim 3, further comprising separating the pluralityof types of charged species into sections.
 7. The method according toclaim 6, wherein the separating of the plurality of types of chargedspecies into sections is based on capillary electrophoresis.
 8. Themethod according to claim 1, wherein the determining of the at least oneproperty of the components of the sample comprises determining aconcentration of at least one of the plurality of types of chargedspecies.
 9. The method according to claim 1, wherein the determining ofthe at least one property of the components of the sample comprisesdetermining a property of a first one of the plurality of types ofcharged species relative to a property of a second one of the pluralityof types of charged species to determine the concentration of any one ofthe plurality of types of charged species.
 10. The method according toclaim 3, wherein the plurality of types of charged species compriseslithium cations.
 11. A disposable apparatus for conducting ameasurement, the disposable apparatus comprising: at least one channeladapted to be filled with a background solution prior to use, the atleast one channel comprising at least one first opening and the at leastone channel being otherwise sealed, a partly permeable layer placedabove the at least one first opening, the partly permeable layercomprising one or more holes permeable to components of a sample, andthe one or more holes being adapted to having the sample placed thereon,and a cover layer placed above the one or more openings, the cover layerbeing adapted to seal the at least one channel from environmentalexposure and to being removed from the one or more openings.
 12. Thedisposable apparatus according to claim 11, wherein the partly permeablelayer is impermeable to at least some components of cellular material.13. The disposable apparatus according to claim 11, wherein the partlypermeable layer is a first layer comprising the at least one firstopening.
 14. The disposable apparatus according to claim 11, furthercomprising at least one first conductivity electrode and at least onesecond conductivity electrode arranged in or nearby the at least onechannel at a distance from the first opening.
 15. The disposableapparatus according to claim 11, wherein the at least one channelcomprises a first reservoir at a first end of the at least one channeland a second reservoir at a second end of the at least one channel. 16.The disposable apparatus according to claim 11, further comprising atleast one first electrophoresis electrode and at least one secondelectrophoresis electrode arranged in or nearby the at least onechannel.
 17. The disposable apparatus according to claim 11, wherein theat least one channel comprises at least one second opening, and whereinthe at least one channel is otherwise sealed.
 18. The disposableapparatus according to claim 11, wherein the partly permeable layer is amembrane.
 19. The disposable apparatus according to claim 11, furthercomprising heating elements placed inside or around the at least onechannel or around the disposable apparatus.
 20. The disposable apparatusaccording to claim 11, wherein the at least one channel is filled with abackground solution prior to shipment.